Development of a steady‐state FRET‐based assay to identify inhibitors of the Keap1‐Nrf2 protein–protein interaction

Schaap, Marjolein; Hancock, Rowena; Wilderspin, Andrew F.; Wells, Geoff

doi:10.1002/pro.2384

Cited by 31 publications

(15 citation statements)

References 35 publications

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“…For this purpose, ECFP was fused to the N terminus of STAT1ND or STAT1(1-713), while EYFP was fused to the N terminus of Y3. If ECFP is close to EYFP (within about 100 Å), emission from ECFP at 475 nm, which is generated after excitation of the ECFP chromophore at 435 nm, is decreased, while emission from EYFP at 527 nm is increased (32). Gel filtration analysis showed that ECFP-fused STAT1ND could bind to EYFP-fused Y3 (Fig.…”

Section: Resultsmentioning

confidence: 97%

Structural Basis of the Inhibition of STAT1 Activity by Sendai Virus C Protein

Oda

Matoba

Irie

et al. 2015

J Virol

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Sendai virus (SeV IMPORTANCE Sendai virus, a paramyxovirus that causes respiratory diseases in rodents, possesses the C protein, which inhibits the signal transduction pathways of interferon alpha/beta (IFN-␣/␤) and IFN-␥ by binding to the transcription factor STAT1. In virusinfected cells, phosphorylation of STAT1 at the Tyr701 residue is potently enhanced, although transcription by STAT1 is inert. Here, we determined the crystal structure of the N-terminal domain of STAT1 associated with the C-terminal half of the C protein. Molecular modeling and experiments suggested that the two C proteins bind to and stabilize the parallel form of the STAT1 dimer, which are likely to be phosphorylated at Tyr 701 , further inducing high-molecular-weight complex formation and inhibition of transcription by IFN-␥. We also discuss the possible mechanism of inhibition of the IFN-␣/␤ pathways by the C protein. This is the first structural report of the C protein, suggesting a mechanism of evasion of the paramyxovirus from innate immunity.

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Section: Resultsmentioning

confidence: 97%

Structural Basis of the Inhibition of STAT1 Activity by Sendai Virus C Protein

Oda

Matoba

Irie

et al. 2015

J Virol

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“…Numerous cell‐based and in silico screens have identified Nrf2‐activating compounds (Schaap, Hancock, Wilderspin, & Wells, 2013; Wang et al, 2013; Williamson et al, 2012; Wu, McDonald, Liu, Chaguturu, & Klaassen, 2012), including triterpenoid 2‐cyano‐3,12‐dioxooleana‐1,9(11)‐dien‐28‐oate‐methylamide (CDDO‐MA), puerarin, sulforaphane, CDDO‐ethyl amide and others. Nrf2 activators demonstrated activity in in vitro and in vivo in different neurodegenerative mouse models, protecting neurons, decreasing the accumulation of aberrant proteins and increasing life span (Buendia et al, 2016; Joshi and Johnson 2012).…”

Section: Potential Therapeutic Targets In Astrocytesmentioning

confidence: 99%

Astroglia as a cellular target for neuroprotection and treatment of neuro‐psychiatric disorders

Liu

Teschemacher

Kasparov

2017

Glia

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Astrocytes are key homeostatic cells of the central nervous system. They cooperate with neurons at several levels, including ion and water homeostasis, chemical signal transmission, blood flow regulation, immune and oxidative stress defense, supply of metabolites and neurogenesis. Astroglia is also important for viability and maturation of stem‐cell derived neurons. Neurons critically depend on intrinsic protective and supportive properties of astrocytes. Conversely, all forms of pathogenic stimuli which disturb astrocytic functions compromise neuronal functionality and viability. Support of neuroprotective functions of astrocytes is thus an important strategy for enhancing neuronal survival and improving outcomes in disease states. In this review, we first briefly examine how astrocytic dysfunction contributes to major neurological disorders, which are traditionally associated with malfunctioning of processes residing in neurons. Possible molecular entities within astrocytes that could underpin the cause, initiation and/or progression of various disorders are outlined. In the second section, we explore opportunities enhancing neuroprotective function of astroglia. We consider targeting astrocyte‐specific molecular pathways which are involved in neuroprotection or could be expected to have a therapeutic value. Examples of those are oxidative stress defense mechanisms, glutamate uptake, purinergic signaling, water and ion homeostasis, connexin gap junctions, neurotrophic factors and the Nrf2‐ARE pathway. We propose that enhancing the neuroprotective capacity of astrocytes is a viable strategy for improving brain resilience and developing new therapeutic approaches.

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“…Additionally, L75, L84 and D77 have been suggested to stabilise the beta‐turn structure. The binding of isolated, unconstrained peptides based on the ETGE motif of Nrf2 to Keap1 is well‐characterized through biophysical, crystallographic and cell‐based assays . Inhibiting this interaction has therapeutic potential for a range of diseases, such as the chemoprevention of cancer, diabetes Alzheimer's disease, and chronic obstructive pulmonary disease …”

Section: Introductionmentioning

confidence: 99%

Exploring new strategies for grafting binding peptides onto protein loops using a consensus‐designed tetratricopeptide repeat scaffold

2019

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Peptide display approaches, in which peptide epitopes of known binding activities are grafted onto stable protein scaffolds, have been developed to constrain the peptide in its bioactive conformation and to enhance its stability. However, peptide grafting can be a lengthy process requiring extensive computational modeling and/or optimisation by directed evolution techniques. In this study, we show that ultra-stable consensus-designed tetratricopeptide repeat (CTPR) proteins are amenable to the grafting of peptides that bind the Kelch-like ECH-associated protein 1 (Keap1) onto the loop between adjacent repeats. We explore simple strategies to optimize the grafting process and show that modest improvements in Keap1-binding affinity can be obtained by changing the composition of the linker sequence flanking either side of the binding peptide.

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Development of a steady‐state FRET‐based assay to identify inhibitors of the Keap1‐Nrf2 protein–protein interaction

Cited by 31 publications

References 35 publications

Structural Basis of the Inhibition of STAT1 Activity by Sendai Virus C Protein

Structural Basis of the Inhibition of STAT1 Activity by Sendai Virus C Protein

Astroglia as a cellular target for neuroprotection and treatment of neuro‐psychiatric disorders

Exploring new strategies for grafting binding peptides onto protein loops using a consensus‐designed tetratricopeptide repeat scaffold

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